Vector biology-Using a mosquito pathogen as a delivery system for anti-malarial a

描述（由申请人提供）：世界上大约46%的人口生活在蚊子传播的疾病，包括疟疾，丝虫病，病毒性脑炎，登革热和黄热病流行的地区。最近确定，昆虫病原真菌金龟子绿僵菌具有在城市环境中控制成蚊的潜力，但前提是其效力增加。我们的目标是生产一种或多种真菌产品，这些产品可以减少按蚊和疟原虫种群，从而显著降低疟疾流行率。我们已经证明，M。金龟子绿僵菌是昆虫选择性蝎毒素AaIT的非常有效的递送系统，并且表达AaIT的菌株产生针对蚊子的有效孢子剂量的9倍减少。这是非常重要的，但杀死时间仍然太慢，无法提供足够的保护。在这个应用程序中，我们提出了实验来比较优化M的几种策略。绿僵菌减少疾病传播的能力。感染真菌的蚊子唾液腺上的子孢子数量显著减少，但其机制尚不清楚。我们将对疟原虫、蚊子和M.绿僵菌这将包括检测M的减毒株。激发超免疫反应的绿僵菌，以确定是否M.绿僵菌可用于使蚊子免受疟原虫感染。此外，我们将比较感染M的蚊子的死亡率和子孢子流行率。表达杀虫蛋白和抗疟原虫蛋白的不同组合的绿僵菌菌株。将确定这些措施是否可以协同使用，以有效减少传播潜力。基于这些结果，我们还将测试使用M.这些基因是不同活性的杂交体，并且可以例如靶向昆虫和疟原虫两者。目前的提案将：1）探索蚊子的免疫系统; 2）开发有可能大大降低疟疾流行率的工具和基因工程真菌; 3）开发M.绿僵菌作为一个易处理的模型系统，可用于筛选新的效应。我们设想在筛选后，通过在M.绿僵菌和/或替代病原体、寄生虫或通过转基因蚊子。该项目旨在设计、构建和评估针对成年蚊子和疟原虫的重组真菌病原体。产生优化的真菌病原体的最重要的可能结果将是由于中断目标寄生虫的传播而减少人类疾病。

DESCRIPTION (provided by applicant): Around 46% of the World's population lives in areas where mosquito-borne diseases including malaria, filariasis, viral encephalitides, dengue and yellow fever are endemic. It was recently established that the insect pathogenic fungus Metarhizium anisopliae has the potential to control adult mosquitoes in an urban setting, but only if its potency is increased. Our goal is to produce one or more fungal products which can deplete anopheline and Plasmodium populations to the extent that marked reductions in malaria prevalence are achieved. We have already shown that M. anisopliae is a very effective delivery system for the insect-selective scorpion toxin AaIT, and that expressing AaIT produced a 9-fold reduction in effective spore doses against mosquitoes. This was very significant but kill times remain too slow for adequate protection. In this application, we propose experiments to compare several strategies for optimizing M. anisopliae's ability to curtail disease transmission. Mosquitoes infected with fungi showed a significant reduction in the number of sporozoites on salivary glands, but the mechanism responsible is unknown. We will carry out a detailed analysis of the interactions between Plasmodium, mosquitoes and M. anisopliae. This will include testing an attenuated strain of M. anisopliae that elicits a hyperimmune response to determine whether M. anisopliae can be used to immunize mosquitoes from Plasmodium. Further, we will compare mortality and sporozoite prevalence in mosquitoes infected with M. anisopliae strains expressing different combinations of insecticidal and anti-plasmodial proteins. It will be determined if these can be used synergistically to achieve effective reductions in transmission potential. Based on these results, we will also test the efficacy of using M. anisopliae to express synthetic multifunctional genes that are hybrids of different activities and that could, for example, target both the insect and the Plasmodium. The current proposal will: 1) explore the mosquito immune system; 2) develop tools and genetically engineered fungi that have the potential to greatly reduce malaria prevalence, and 3) develop M. anisopliae as a tractable model system that can be used to screen novel effectors. We envisage that after screening, the most potent effectors could be delivered against mosquitoes or Plasmodium by expression in M. anisopliae and/or in alternative pathogens, commensals or via transgenic mosquitoes. PUBLIC HEALTH RELEVANCE This project aims to design, construct and evaluate recombinant fungal pathogens that target adult mosquitoes and the malaria parasite. The most significant possible outcome of producing an optimized fungal pathogen will be a reduction of human disease as a result of interrupting transmission of the target parasite.